Beta-xylosidase (1,4-.beta.-D-xylan-xylohydrolase; EC 3.2.1.37) is one of the xylanolytic enzymes. Xylans are a major constituent of the cell walls of plants, and are second only to cellulose. They are abundantly found in most land plants, especially in agricultural by-products such as straw, wheat-bran, corn cobs, cotton seed, etc. Xylan is a complex polymer consisting of a .beta.-1,4-linked xylose polymer with arabino-furanose, glucuronic acid, methylglucuronic acid and acetyl side-groups. Endoxylanase (EC 3.2.1.8) randomly cleaves the .beta.-1,4-bonds in the xylan backbone to yield oligosaccharides, xylobiose and xylose. Beta-xylosidase cleaves terminal xylose units from the non-reducing end of the xylose oligomers resulting from endoxylanase activity. .alpha.-Glucuronidase cleaves glucuronic acid side groups from backbone xylose units, whereas .alpha.-L-arabinofuranosidases (EC 3.2.1.55) cleave arabinose units from the xylan backbone and acetylesterases (EC 3.1.1.6) remove the acetyl side-groups.
Beta-xylosidase is also effective in transglycosylation reactions wherein monosaccharide units or alcohols are attached to or cleaved from xylose units. Beta-xylosidase is rate-limiting in xylan hydrolysis (Dekker 1983, Poutanen and Puls 1988).
The hydrolysing and transglycosylating reactions of .beta.-xylosidases are economically important for the breakdown and utilisation of agricultural waste material e.g. in the production of xylose, xylose oligomers and xylitol, which are useful as sweeteners in foodstuffs, candies and medicaments, especially as a sugar substitute. Also, the enzyme or its products can be used as bread improvers and in the beer brewing industry.
Beta-xylosidases have been isolated from various sources including bacteria and fungi. For example, the purification of .beta.-xylosidase from Aspergillus niger was reported by Rodionova et al. (1983); the molecular weight was reported to be 253,000 on the basis of gel filtration and 122,000 on the basis of SDS electrophoresis, whereas its isoelectric point was at pH 4.9.
Three endoxylanases and one .beta.-xylosidase were isolated from Aspergillus awamori by Kormelink et al. (1993); the .beta.-xylosidase had a molecular weight of 110,000, a pH optimum of 6.5 and a temperature optimum of 70.degree. C.
Beta-D-xylosidase from rumen fungus Neocallirnastix frontalis was reported by Garcia-Campayo and Wood (1993) and had an apparent molecular weight (gel filtration) of 150,000, a pH optimum of 6.4 and a temperature optimum of 37.degree. C. Utt et al (1991) report the sequencing of the xylB of the ruminal bacterium Butyrivibrio fibrisolvens encoding both .beta.-xylosidase and .alpha.-arabinofuranosidase activities.
Known .beta.-xylosidases have activity patterns that do not always correspond to the industrial needs. In particular it is often desirable that the enzyme has a high xylosidase specificity and low specificities for other substrates, such as glucosides and galactosides. Especially fungal .beta.-xylosidases are highly advantageous for their activity levels and specificity patterns.
In order to be able to provide .beta.-xylosidase-like enzymes having the desired activity patterns from the desired production organisms, sequence information of the .beta.-xylosidase gene should be available. Up to now however, no sequence information on fungal .beta.-xylosidases has been reported.